Recurrent infection and deterioration of lung function are the major causes of morbidity and mortality in cystic fibrosis (CF). Although the genetic defect in CF was discovered in 1989, the mechanisms by which CFTR mutations cause lung disease in CF remain uncertain. This proposal focuses on the role of the airway surface liquid (ASL) and airway submucosal glands in CF lung disease. Our laboratory recently developed a series of novel fluorescent indicators and microscopy methods to measure ASL depth and composition, and submucosal gland secretion rate, fluid composition and viscosity. Aim 1 will determine whether ASL composition is abnormal in CF, focusing on lower airway function, clinically relevant stresses, and utilizing human tissues. Aim 2 will determine whether submucosal gland function is defective in CF, focusing on tissues that have not been subject to chronic infection and inflammation. Aim 3 will examine the hyper-absorption hypothesis (thinned, hyperviscous ASL in CF) and the defective oxygenation hypothesis (hypoxic ASL in CF). Some of the measurements will be done using intact human airways (normal vs. CF), including bronchoscopic biopsies obtained from pediatric CF patients before significant airway disease has developed. Some experiments will utilize potent small-molecule CFFR inhibitors and activators recently developed by our lab. The novel approaches and model systems should provide definitive data on putative abnormalities in ASL composition and submucosal gland function in CF. Understanding of lung disease mechanisms is of central importance in CF research, having direct implications for development of evaluation of new therapies. [unreadable] [unreadable]